The objective of this project is the identification of specific Alzheimer's disease (AD)-associated proteases that will serve as early diagnostic markers for AD, and potentially as therapeutic targets for drug design and discovery effort outlined within the overall DDG-AD Program Project. Today, accurate diagnosis of AD can be made only after pathological examination of autopsy brain tissue that has been stained, or more recently through the use of Abbott's Alz-EIA test which measures brain tissue levels of Alzheimer's disease associated protein (ADAP). As research progresses towards development of effective, rationally designed therapeutics, it will be essential to develop early, accurate diagnostics for AD. The specific aims that constitute this overall objective are: 1) To identify AD- specific protease markers in human AD brain tissue, and provide purified fractions containing protease markers to Project 3 final purification; 2) To develop antibodies, diagnostic immunoassays and diagnostic protease activity assays for A specific protease markers; 3) To establish a clinical correlation between putative AD-associated protease markers are AD; 4) To study the distribution of AD-associated protease markers in various brain regions and in CSF, blood, urine and other tissues, and in brain tissue from transgenic mice; 5) To determine the relationship and/or association of A associated protease markers with other biochemical markers of AD and neural damage; 6) To prepare specific antibodies and protein fractions for APP processing studies (with Project 4), cell biology studies of APP catabolism (with Project 5), NGFR processing (with Project 6) and for neuropharmacologics studies (with Core F). The method and procedures deployed to accomplish these aims include protein biochemical, analytical biochemical, immunological, immunohistochemical and molecular biological techniques. Collaborations with other projects include: Project 1- identification of AD-specific proteases and development of diagnostic tests based on novel peptide probes; Project purification of AD-associated proteases that will be used in structure based drug design by Project 1; Project 4-generation of unique antibody reagents and reassessments of AD markers in transgenic animals developed in Project 4 with the Transgenic Center and Northwestern University; Project 5-studies with protease- containing tissue fraction and specific antibody reagents in neuronal cell culture; Project 6-studies to establish the utility of truncated NGF receptor as a diagnostic marker and to evaluate effects of AD-protease fractions on NGF receptor processing in cell culture. Within this multi- disciplinary drug discovery program project, these studies will provide the ultimate clinical correlation between any potential protease drug target for AD therapy and authentic relevance to the Alzheimer's disease process. Any AD drug candidate developed in this DDG-AD that acts by blocking an early causative event of Ad will be useless if administered to a patient population identified only be late stage cognitive deficits that undoubtedly can be reversed by such a drug. Thus, at the onset of a major drug discovery effort such as this, it is crucial to embark an intensive parallel path towards development of early, accurate diagnostic assays.